Expression Frequency of the Human Immunoglobulin
 Light Chain Genes and the Roles of Their
Recombination Signal and Spacer Sequences
Taku Okada 12
Masaru Tomita 13
dachan@sfc.keio.ac.jp
mt@sfc.keio.ac.jp
4Kazuhiko Kawasaki
Shinsei Minoshima 4
Nobuyoshi Shimizu 4

kawasaki@dmb.med.keio.ac.jp

mino@dmb.med.keio.ac.jp

shimizu@dmb.med.keio.ac.jp

1 Laboratory for Bioinformatics
2 Graduate School of Media and Governance
3 Department of Environmental Information

Keio University, 5322 Endo, Fujisawa, 252-8520, Japan

4 Department of Molecular Biology, Keio University School of medicine
35 Shinanomachi, Shinjuku, Tokyo, Japan

1 Introduction
The recombination signal sequences (RSS) is the essential element for the immunoglobulin gene rearrangement. The RSS consists of 3 elements: heptamer (7mer), 12-bp or 23-bp spacer, and nonamer(9mer). The heptamer and nonamer are well conserved [6]. The rearrangement occurs between anRSS with 12 bp spacer sequences and an RSS with 23 bp spacer sequences (the `12/23 rule') [1]. RSSsof indispensable V gene segments should be well conserved to maintain high eciency of the V(D)Jjoining.
In this work, the RSSs of the human immunoglobulin  light chain genes were investigated how therecombination and expression frequencies of the genes are in uenced by the RSSs. We rst conducted
an extensive database search to estimate expression frequencies of the variable region gene for the light chain (V ) combination with the joining (J) and constant (C) region.
The genes for the  light chain genes located on chromosome 22q11 have been sequenced recently
by two di erent groups [4, 7]. In this one-megabase long locus, Kawasaki et al. [4] reported that there
are 36 functional V gene segments and 33 pseudogene segments, followed by 7 J C gene segments,four of which are functional. On the other hand, Williams et al. reported that there are 37 V genesegments, 30 of which are functional. The di erences between these two reports are probably due toallelic variations.

2 Results
Fig. 1 shows the important nucleotides of RSS of the V gene segment. The 1st, 2nd, 3rd, and 7th
nucleotides in the heptamer and the 1st, 6th, 7th, and 8th nucleotides in the nonamer are highly
conserved among the expressed segments, indicating that those bases are crucial for recombinationeciency, which is consistent with previously reported results of laboratory experiments [2, 3, 5]. Unexpectedly, however, the 15th, 16th, 17th, and 18th nucleotides in the 23bp spacer are well conserved,
suggesting that the spacer sequence also plays an important role in the recombination process in the light chain gene.

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heptamer
3
4
5

6

7

1

2

3

4

5

spacer
6
7
8

9

10

11

12

13

T

G

A

T

C

C

A

GGCCCATG

16spacer
17 18 1920

21

22

23

1

2

3nonamer
45

6789GATGAGACAAAACC

12CA

CA

1415

GGGA

C

The nucleotide is present in morethan 99% of the expressed segments

AThe nucleotide is present in morethan 95% of the expressed segments

Figure 1: The conservation of the RSS of V gene segments.

3 Conclusion
We have shown that, besides the heptamer and the nonamer, the spacer sequence also plays animportant role in the VJ joining of human immunoglobulin  genes. Speci cally, it is suggested thatthe second half of the 23-bp spacer sequence a ect joining eciency because the nucleotide sequenceof this region is well conserved among all RSSs, and because the V segments substituted at one ofthese nucleotides are expressed in signi cantly lower frequencies.

References[1] Bogue, M. and Roth, D.B., Mechanism of V(D)J recombination, Curr. Opin. Immunol., 8:175{180, 1996.
[2] Cuomo, C.A., Mundy, C.L. and Oettinger, M.A., DNA Sequence and Structure Requirements forCleavage of V(D)J Recombination Signal Sequences, Mol. Cell. Biol., 16:5683{5690, 1996.
[3] Hesse, J.E., Lieber, M.R., Mizuuchi, K., and Gellert, M., V(D)J recombination: a functionalde nition of the joining signals, Genes Dev., 3:1053{1061, 1989.
[4] Kawasaki, K., Minoshima, S., Nakato, E., Shibuya, K., Shintani, A., Schmeits, J.L., Wang, J.,
and Shimizu, N., One-Megabase Sequence Analysis of the Human Immunoglobulin  Gene Locus,
Genome Res., 7:250{261, 1997.
[5] Ramsden, D.A., McBlane, J.F., van Gent, D.C., and Gellert, M., Distinct DNA sequence andstructure requirements for the two steps of V(D)J recombination signal cleavage, Embo J.,
15:3197{3206, 1996.
[6] Tonegawa, S., Somatic generation of antibody diversity, Nature, 302:575{581, 1983.
[7] Williams, S.C., Frippiat, J.P., Tomlinson, I.M., Ignatovich, O., Lefranc, M.P., and Winter, G.,
Sequence and Evolution of the Human Germline V Repertoire, J. Mol. Biol., 264:220{232, 1996.

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